Human hepatitis B virus (HBV) is a cause of both acute and chronic liver disease in man, and chronic infections are epidemiologically associated with hepatocellular carcinoma. HBV is the prototype of a new family of hepatotropic DNA viruses (hepadna viruses) which includes woodchuck hepatitis virus, ground squirrel hepatitis virus, and duck hepatitis B virus (DHBV). Our recent studies with DHBV indicate a novel mode of replication for the hepadna viruses, involving reverse transcription of an RNA intermediate occurring as a step in virion maturation. The long term objective in studying the hepadna viruses is to understand and prevent the phenomenon of viral pathogenesis. An important aspect of this problem is to understand the molecular biology of virus replication, including the mechanism of initiation of infection and the control of virus expression. We have been studying these processes using DHBV as a model of human hepatitis B virus. During the next three years, our main objectives will be in two areas. First, we propose to characterize in detail the initiation of infection in embryos and ducklings, and the basis for the age dependent decline in susceptibility. Second, we will continue work on the mechanism of virus DNA synthesis, including the structure of the RNA template for minus strand synthesis and the DNA template for plus strand synthesis, and the mechanisms and timing for initiation and elongation of both strands. As a new area of research in our lab, we will also carry out a pilot project on the induction and role of liver specific immunoregulatory factors in viral hepatitis in Pekin ducks. Towards these goals, we have demonstrated that infection is rapid (less than 24h) in embryos and young ducklings, allowing a detailed characterization of early steps in infection without recourse to tissue culture systems, which are unavailable. In addition, DNA synthesis complexes are readily isolated from the liver, permitting analysis of virion DNA synthesis in an in vitro system. Cloned DNA as well as antibodies have been prepared to assist both studies, and we have been able to use the former to identify the origin and a potential primer (protein) of reverse transcription.